Humans are increasingly exposed to ionizing radiation as a result of rapidly expanding volume of diagnostic and therapeutic radiation, nuclear power accidents such as Chernobyl and Fukushima, high altitude travel, and other exposures. Exposure to radiation is known to increase the risk of various cancers including thyroid cancer. However, the molecular mechanisms of radiation-induced carcinogenesis remain poorly understood. During the previous cycles of this proposal, we have established the central role of chromosomal rearrangements, such as RET/PTC, in radiation-induced thyroid carcinogenesis, and created in vitro models of dose-dependent induction of RET/PTC in human thyroid cells by ?-radiation. Moreover, we have also obtained and genotyped 70 post-Chernobyl thyroid tumors from patients with carefully reconstructed thyroid dose received from 131I and identified 20 tumors associated with high 131I dose that were negative for all known mutations. Our recent analysis of this cohort revealed a strong link between RET/PTC and leaving in the regions of iodine deficiency, which we will explore in this proposal to study the reasons for the association between iodine deficiency and cancer risk found after Chernobyl. Moreover, our first RNA-Seq run of one of the mutation-negative tumors associated with high 131I dose led to the discovery of a novel chromosomal rearrangement, which we find to be the second most common type of chromosomal rearrangements in post- Chernobyl cancers after RET/PTC. These valuable tools will be used in the current proposal, which will continue to dissect the mechanisms of chromosomal rearrangements and radiation carcinogenesis in the thyroid. Specifically, we will test the hypothesis that the rate of generation of RET/PTC rearrangements by radiation in thyroid cells is influenced by cell cycle stage at the time of exposure and transcriptional status of genes in the regions undergoing recombination. We will also determine whether downregulation of ATM and other homologous recombination repair genes enhances RET/PTC induction by radiation in thyroid cells in vitro, and if these genes are involved in the individual susceptibility to radiation carcinogenesis in humans. Finally, we will continue using new sequencing technologies to identify novel types of chromosomal rearrangements occurring in thyroid cancer associated with high radiation dose to the thyroid, and will test if the newly identified genetic events can be induced in human thyroid cells by in vitro radiation. These studies will expand our understanding of the genetic mechanisms of radiation-induced thyroid cancer and provide novel information that can be used to identify those individuals who are most susceptible to radiation carcinogenesis and to develop measures for better protection of human populations against the carcinogenic effects of ionizing radiation in a variety of settings such as medical therapeutic radiation, occupational radiation exposure, and nuclear power accidents and nuclear terrorism.